Aptamer-modified carbon nanomaterial based sorption coupled to paper spray ion mobility spectrometry for highly sensitive and selective determination of methamphetamine.

A cellulose paper was modified with an aptamer against methamphetamine on either carbon dots (CDs) or on multichannel carbon nanotubes (CNTs). The resulting sorbent was applied to the extraction of METH from blood or saliva. The METH-loaded paper than also was directly applied as a paper spray ionization source in ion mobility spectrometry. The carbon nanomaterial enhances sensitivity, and the aptamer enhances selectivity. The materials were covalently bound to the paper on one side, while the aptamer was immobilized on the other. After optimization of the extraction process and instrumental parameters, the limits of detection when using the aptamer-CNT modified paper are 0.6 ng·mL-1 for saliva, and 0.45 ng·mL-1 for plasma. The respective values when using aptamer-CD modified paper are 1.5 ng·mL-1 for saliva and 0.9 ng·mL-1 for plasma. Calibration plots are linear in the 2 to 150 ng·mL-1 METH concentration range for saliva, and in the 1.5 to 200 ng·mL-1 concentration ranges for blood when using the aptamer-CNT based method. When using the aptamer-CDs, the dynamic ranges extend from 5 to 200 ng·mL-1 and from 3 to 250 ng·mL-1, respectively. The method was applied to the determination of METH in real samples of saliva and blood, and the accuracy of the method was confirmed by comparison of the results with data analyzed by GC-MS. Graphical abstract ᅟ.

Rapid screening of methamphetamines in human serum by headspace solid-phase microextraction using a dodecylsulfate-doped polypyrrole film coupled to ion mobility spectrometry.

A simple, rapid and highly sensitive method for simultaneous analysis of methamphetamine (MA) and 3,4-methylenedioxy methamphetamine (MDMA) in human serum was developed using the solid-phase microextraction (SPME) combined with ion mobility spectrometry (IMS). A dodecylsulfate-doped polypyrrole (PPy-DS) was applied as a new fiber for SPME. Electrochemically polymerized PPy is formed on the surface of a platinum wire and will contain charge-compensating anion (dodecylsulfate) incorporated during synthesis using cyclic voltammetry (CV) technique. The extraction properties of the fiber to MA and MDMA were examined, using a headspace-SPME (HS-SPME) device and thermal desorption in injection port of IMS. The results show that PPy-DS as a SPME fiber coating is suitable for the successful extraction of these compounds. This method is suitable for the identification and determination of MAs, is not time-consuming, requires small quantities of sample and does not require any derivatization. Parameters like pH, extraction time, ionic strength, and temperature of the sample were studied and optimized to obtain the best extraction results. The HS-SPME-IMS method provided good repeatability (RSDs<7.8 %) for spiked serum samples. The calibration graphs were linear in the range of 20-4000 ng ml(-1) (R(2)>0.99) and detection limits for MDMA and MA were 5 and 8 ng ml(-1), respectively. HS-SPME-IMS of non-spiked serum sample provided a spectrum without any peak from the matrix, supporting an effective sample clean-up. Finally, the proposed method was applied for analysis one of the ecstasy tablet.

Evaluation of Dräger DrugTest 5000 in a Naturalistic Setting.

Reliable field testing devices for psychoactive drugs would be useful tools for the police for detecting drug-impaired drivers. The Norwegian Mobile Police Service (NMPS) started using Dräger DrugTest 5000 (DDT5000) in 2015 as an on-site screening instrument for drugs in samples of oral fluid. The aim of this study was to compare the results of field testing of DDT5000 with drug findings in blood and oral fluid samples taken from drivers suspected for driving under the influence of drugs (DUID). In total, 369 drivers were included in this field testing; blood samples were obtained from all of them, while oral fluid samples were collected with the Intercept device from 301 of them. The median time from field testing with DDT5000 and collection of blood and oral fluid samples was 50 min. The proportions of false positive results with DDT5000 compared to findings in blood samples above the Norwegian legal per se limits were for cannabis 14.5%, amphetamine 23.2%, methamphetamine 38.4%, cocaine 87.1%, opiates 65.9% and benzodiazepines 36.4%. The proportions of false negatives were for cannabis 13.4%, amphetamine 4.9%, methamphetamine 6.1%, cocaine 0.0%, opiates 0.0% and benzodiazepines 18.8%. Among drivers who had drug concentrations above the legal limits in blood, the proportion who tested positive using DDT5000 was 82.9% for THC, 90.8% for amphetamine, 75.7% for methamphetamine, 100.0% for cocaine, 100.0% for opiates and 37.2% for benzodiazepines. In cases with false-positive DDT5000 results compared to blood, traces of drugs were most often found in oral fluid. The DDT5000 did not absolutely correctly identify DUID offenders due to fairly large proportions of false-positive or false-negative results compared to drug concentrations in blood. The police reported that DDT5000 was still a valuable tool in identifying possible DUID offenders, resulting in more than doubling the number of apprehended DUID offenders.

Methamphetamine disposition in oral fluid, plasma, and urine.

This review of the disposition of methamphetamine in oral fluid, plasma, and urine is based on a comprehensive controlled dosing study involving five healthy, drug-free research volunteers who resided on a closed clinical ward for 12 weeks. Subjects were administered four low (10 mg) and high (20 mg) daily oral doses of methamphetamine in two separate sessions. Near-simultaneous collections of oral fluid and plasma were performed on the first day of each low- and high-dose session. Thereafter, oral fluid was provided on each day of dosing by different oral fluid collection methods. All urine specimens were collected on an ad libitum basis throughout the study. Specimens were analyzed by gas-chromatography mass spectrometry for methamphetamine and the metabolite, amphetamine, with a limit of quantification of 2.5 ng/mL for each analyte. Methamphetamine and metabolite concentrations in oral fluid appeared to follow a similar time course in oral fluid as in plasma and were dose-proportional, but oral fluid concentrations exceeded plasma concentrations. Urine drug concentrations were substantially higher than those in oral fluid. Some drug accumulation was noted with daily dosing, but generally did not markedly influence detection times or detection rates of oral fluid tests. Detection times and detection rates for oral fluid and urine were determined at cessation of 4 days of dosing. Generally, detection times and rates for urine were longer than those observed for oral fluid at conventional cutoff concentrations. When contemplating selection of oral fluid as a test matrix, the advantages of oral fluid collection should be weighed against its shorter time of detection compared to that of urine.

Long-circulating liposomes radiolabeled with [18F]fluorodipalmitin ([18F]FDP).

Synthesis of a radiolabeled diglyceride, 3-[(18)F]fluoro-1,2-dipalmitoylglycerol [[(18)F]fluorodipalmitin ([(18)F]FDP)], and its potential as a reagent for radiolabeling long-circulating liposomes were investigated. The incorporation of (18)F into the lipid molecule was accomplished by nucleophilic substitution of the p-toluenesulfonyl moiety with a decay-corrected yield of 43+/-10% (n=12). Radiolabeled, long-circulating polyethylene-glycol-coated liposomes were prepared using a mixture of 1,2-dipalmitoyl-sn-glycero-3-phosphocholine, cholesterol, 1,2-distearoyl-sn-glycero-3-phosphoethanolamine-N-[methoxy(polyethyleneglycol)-2000] ammonium salt (61:30:9) and [(18)F]FDP with a decay-corrected yield of 70+/-8% (n=4). PET imaging and biodistribution studies were performed with free [(18)F]FDP and liposome-incorporated [(18)F]FDP. Freely injected [(18)F]FDP had the highest uptake in the liver, spleen and lungs. Liposomal [(18)F]FDP remained in blood circulation at near-constant levels for at least 90 min, with a peak concentration near 2.5%ID/cc. Since [(18)F]FDP was incorporated into the phospholipid bilayer, it could potentially be used for radiolabeling a variety of lipid-based drug carriers.

Polypeptide with electroactive endgroups as sensing platform for the abused drug 'methamphetamine' by bioelectrochemical method.

Affinity-type sensors have emerged as outstanding platforms in the detection of diagnostic protein markers, nucleic acids and drugs. Thus, these novel platforms containing antibodies could be integrated into the monitoring systems for abused drugs. Herein, we established a novel detection platform for the analysis of a common illicit drug; methamphetamine (METH). Initially, a fluorescent-labeled polypeptide (EDOT-BTDA-Pala), derived from L-alanine N-carboxyanhydride (L-Ala-NCA) via ring-opening polymerization using 4,7-bis(2,3-dihydrothieno[3,4-b][1,4]dioxin-5-yl)benzo[c][1,2,5]thiadiazole-5,6-diamine (EDOT-NH2-BTDA) as initiator, was employed as a glassy carbon electrode (GCE) covering host, in order to immobilize the METH-selective antibody. Prior to the examination of analytical features, GCE/EDOT-BTDA-Pala/Antibody surface was successfully characterized in the way of electrochemical (cyclic voltammetry and electrochemical impedance spectroscopy) and microscopic techniques (scanning electron microscopy and fluorescence microscopy). As for the analytical characterization, linearity and limit of detection (LOD) were found as 10-100µg/mL with an equation of y=0.0429x-0.2347, (R2=0.996) and 13.07µg/mL, respectively. Moreover, sample application using artificial urine, saliva and serum samples spiked with METH (10, 25, 50µg/mL) were performed and LC-MS/MS system was used for further confirmation. The described platform can be adapted to monitor the other types of abused drugs by using suitably selected biorecognition elements.

Development and evaluation of electromembrane extraction across a hollow polymer inclusion membrane.

In this work, a new variation of the electromembrane extraction (EME) approach employing a hollow polymer inclusion membrane (HPIM) was developed. In this method, a HPIM was prepared by casting a solution of the desired proportions of cellulose acetate (CTA), tris(2-ethylhexyl)phosphate (TEHP) and di-(2-ethylhexyl)phosphoric acid (D2EHPA) in dichloromethane on glass capillary tubing. Three basic drugs namely amphetamine, methamphetamine, and 3,4-methylenedioxy-N-methylamphetamine (MDMA) were selected as model analytes to evaluate the extraction performance of this new approach. The drugs were extracted from human plasma samples, through a 20µm thickness HPIM, to an aqueous acceptor solution inside the lumen of the hollow membrane. Parameters affecting the extraction efficiency were investigated in detail. Under the optimized conditions, enrichment factors in the range of 97-103-fold were obtained from 3mL of sample solution with a 10min extraction time and an applied voltage of 300V across the HPIM. The detection limits of the method for the three drugs were in the range of 1.0-2.5ng/mL (at a signal/noise ratio of three), with relative standard deviations of between 6.4% and 7.9%. When the method was applied to spiked plasma samples, the relative recoveries ranged from 99.2% to 100.8%. Enrichment factors of 103, 99 and 97 were obtained for amphetamine, methamphetamine, and MDMA, respectively. A comparison was also made between the newly developed approach and EME using supported liquid membranes (SLM) as well as standard sample preparation methods (liquid-liquid extraction) used by the Toxicology Unit, Department of Chemistry, Malaysia.

Oral fluid with three modes of collection and plasma methamphetamine and amphetamine enantiomer concentrations after controlled intranasal l-methamphetamine administration.

Methamphetamine is included in drug testing programmes due to its high abuse potential. d-Methamphetamine is a scheduled potent central nervous system stimulant, while l-methamphetamine is the unscheduled active ingredient in the over-the-counter nasal decongestant Vicks® VapoInhaler™. No data are available in oral fluid (OF) and few in plasma after controlled Vicks® VapoInhaler™ administration. We quantified methamphetamine and amphetamine enantiomers in OF collected with two different devices and plasma via a fully validated liquid chromatography-tandem mass spectrometry (LC-MS/MS) method. Additionally, OF were analyzed with an on-site screening device. Sixteen participants received 7 Vicks® VapoInhaler™ doses according to manufacturer's recommendations. Specimens were collected before and up to 32 h after the first dose. No d-methamphetamine or d-amphetamine was detected in any sample. All participants had measurable OF l-methamphetamine with median maximum concentrations 14.8 and 16.1 µg/L in Quantisal™ and Oral-Eze® devices, respectively, after a median of 5 doses. One participant had measurable OF l-amphetamine with maximum concentrations 3.7 and 5.5 µg/L after 6 doses with the Quantisal™ and Oral-Eze® devices, respectively. There were no positive DrugTest® 5000 results. In the cutoff range 20-50 µg/L methamphetamine with amphetamine ≥limit of detection, 3.1-10.1% of specimens were positive; first positive results were observed after 1-4 doses. Two participants had detectable plasma l-methamphetamine, with maximum observed concentrations 6.3 and 10.0 µg/L after 2 and 5 doses, respectively. Positive OF and plasma methamphetamine results are possible after Vicks® VapoInhaler™ administration. Chiral confirmatory analyses are necessary to rule out VapoInhaler™ intake. Implementing a selective d-methamphetamine screening assay can help eliminate false-positive OF results.

A Nanotechnology-Based Platform for Extending the Pharmacokinetic and Binding Properties of Anti-methamphetamine Antibody Fragments.

To address the need for effective medications to aid in the treatment of methamphetamine (METH) abuse, we used a nanotechnology approach to customize the in vivo behavior of an anti-METH single chain antibody (scFv7F9Cys). Anti-METH scFv7F9Cys was conjugated to dendrimer nanoparticles via a polyethylene glycol (PEG) linker to generate high-order conjugates termed dendribodies. We found that the high affinity (KD = 6.2 nM) and specificity for METH was unchanged after nanoparticle conjugation. The dendribodies were administered in an i.v. bolus to male Sprague Dawley rats after starting a s.c. infusion of METH. The PCKN values for clearance and volume of distribution of scFv7F9Cys after conjugation to dendrimers decreased 45 and 1.6-fold respectively, and the terminal elimination half-life increased 20-fold. Organ distribution of scFv7F9Cys and dendribody in blood and urine agreed well with the PCKN data. Renal clearance appeared to be the major route of elimination for both experimental medications. We have thus successfully developed a novel multivalent METH-binding nanomedicine by conjugating multiple anti-METH scFvs to dendrimer nanoparticles, extending the scFv half-life from 1.3 (± 0.3) to 26 (± 2.6) hr. These data suggest that the dendribody design could be a feasible platform for generating multivalent antibodies with customizable PCKN profiles.

Radioimmunoassay of amphetamines in rat parotid saliva.

Amphetamines, a commonly abused class of drugs, have been detected in various biological specimens, in particular, urine and blood. However, little information is available concerning the detection of these drugs in saliva. This investigation, utilizing the rat salivary secretions, has been attempted to establish the ability of amphetamines to be secreted in saliva and to determine the feasibility of using radioimmunoassay (RIA) for drug detection in saliva. The results of this investigation showed that (1) d-amphetamine and methamphetamine decreased salivary flow, (2) after d-amphetamine RIA tests were demonstrated in both saliva and plasma for a period of fifty minutes, and (3) positive RIA reactions were obtained by the following metamphetamine metabolites: amphetamine, 4-hydroxynorephedrine and 4-hydroxyamphetamine. Methamphetamine and 4-hydroxy-N-methylamphetamine were found to be non-reactive in the radioimmunoassay procedure. The results indicate that saliva could be radioimmunoassayed for the detection of amphetamine or amphetamine derivatives after the administration of either d-amphetamine and methamphetamine.

Development of a rapid and sensitive method for the quantitation of amphetamines in human plasma and oral fluid by LC-MS-MS.

Target analysis of amphetamines in biological samples is of great importance for clinical and forensic toxicologists alike. At present, most laboratories analyze such samples by gas chromatography-mass spectrometry. However, this procedure is labor-intensive and time-consuming, particularly as a preliminary extraction and derivatization are usually unavoidable. Here we describe the development of an alternative method. Amphetamines were isolated from human plasma and oral fluid using a simple methanol precipitation step and subsequently analyzed using reversed-phase liquid chromatography-tandem mass spectrometry. Quantitation of the drugs was performed using multiple reaction monitoring. The developed method, which requires only 50 microL of biological sample, has a total analysis time of less than 20 min (including sample preparation) and enables the simultaneous quantitation of 3,4-methylenedioxymethamphetamine, 3,4-methylenedioxyamphetamine, 3,4-methylenedioxyethylamphetamine, amphetamine, methamphetamine, and ephedrine in a single chromatographic run. Limits of detection of 2 microg/L or better were obtained. The method has been validated and subsequently applied to the analysis of plasma and oral fluid samples collected from current drug users.

Rapid quantitative chiral amphetamines liquid chromatography-tandem mass spectrometry: method in plasma and oral fluid with a cost-effective chiral derivatizing reagent.

Methamphetamine is a widely abused psychostimulant containing a chiral center. Consumption of over-the-counter and prescription medications may yield positive amphetamines results, but chiral separation of l- and d-methamphetamine and its metabolite amphetamine can help determine whether the source was licit or illicit. We present the first LC-MS/MS method with precolumn derivatization for methamphetamine and amphetamine chiral resolution in plasma and oral fluid collected with the Oral-Eze(®) and Quantisal™ devices. To 0.5mL plasma, 0.75mL Oral-Eze, or 1mL Quantisal specimen racemic d11-methamphetamine and amphetamine internal standards were added, followed by protein precipitation. Samples were centrifuged and supernatants loaded onto pre-conditioned Phenomenex(®) Strata™-XC Polymeric Strong Cation solid phase extraction columns. After washing, analytes were eluted with 5% ammonium hydroxide in methanol. The eluate was evaporated to dryness and reconstituted in water. Derivatization was performed with 1-fluoro-2,4-dinitrophenyl-5-l-alanineamide (Marfey's reagent) and heating at 45°C for 1h. Derivatized enantiomer separations were performed under isocratic conditions (methanol:water, 60:40) with a Phenomenex(®) Kinetex(®) 2.6µm C18 column. Analytes were identified and quantified by two MRM transitions and their ratio on a 3200 QTrap (AB Sciex) mass spectrometer in ESI negative mode. In all three matrices, the method was linear for all enantiomers from 1 to 500µg/L, with imprecision and accuracy of ≤11.3% and 85.3-108%, respectively. Extraction efficiencies ranged from 67.4 to 117% and matrix effects from -17.0 to 468%, with variation always ≤19.1%. Authentic plasma and OF specimens were collected from an IRB-approved study that included controlled Vicks(®) VapoInhaler™ administration. The present method is sensitive, selective, economic and rapid (separations accomplished in <10min), and improves methamphetamine result interpretation.

Detection of Delta9-tetrahydrocannabinol and amphetamine-type stimulants in oral fluid using the Rapid Stat point-of-collection drug-testing device.

The Rapid Stat assay, a point-of-collection drug-testing device for detection of amphetamines, cannabinoids, cocaine, opiates, methadone, and benzodiazepines in oral fluid, was evaluated for cannabis and amphetamine-type stimulants. The Rapid Stat tests (n = 134) were applied by police officers in routine traffic checks. Oral fluid and blood samples were analyzed using gas chromatography-mass spectrometry (GC-MS) for Delta(9)-tetrahydrocannabinol, amphetamine, methamphetamine, methylenedioxymethamphetamine, methylenedioxyethylamphetamine, and methylenedioxyamphetamine. The comparison of GC-MS analysis of oral fluid with the Rapid Stat results for cannabis showed a sensitivity of 85%, a specificity of 87%, and a total confirmation rate of 87%. When compared with serum, the sensitivity of the cannabis assay decreased to 71%, the specificity to 60%, and the total confirmation rate to 66%. These findings were possibly caused by an incorrect reading of the THC test results. Comparison of the Rapid Stat amphetamine assay with GC-MS in oral fluid showed a sensitivity of 94%, a specificity of 97%, and a total confirmation rate of 97%. Compared with serum, a sensitivity of 100%, a specificity of 90%, and a total confirmation rate of 92% was found. The amphetamine assay must, therefore, be regarded as satisfactory.

Drugs and driving: the Finnish perspective.

Drugs can cause behavioural impairment of the driver's ability to operate safely That impairment of driving ability can be documented, and biological fluids can be tested for drugs. Most countries have legislation that covers driving under the influence of alcohol and/or drugs. Some countries have introduced zero-tolerance laws (per se laws), which prohibit the operation of a motor vehicle while an illicit drug or its metabolite is present in the body, whether or not impairment is manifested. There is growing interest in using saliva (oral fluid) in preliminary roadside testing. Legislation in the state of Victoria, Australia, already allows the use of oral fluid for evidentiary testing in the case of cannabis and methamphetamine. Nevertheless, blood testing will probably remain the most common form of evidentiary testing. It has been estimated that the prevalence of illicit drug use among the general driving population in Europe is in the range of 1-5 per cent, while the prevalence of licit drugs, such as benzodiazepines, affecting driving performance is higher: 5-10 per cent. Epidemiological research is often carried out on offenders and drivers involved in collisions. Among drivers suspected of driving under the influence of drugs, there is a high percentage of licit and/or illicit drug use, as the statistics for Finland in the present article show. The drugs of most concern are amphetamine and amphetamine-type substances, cocaine, cannabis, opiates and benzodiazepines and other sedative-hypnotics. The handling of drugs and driving cases are presented, and a summary of areas for further study are provided.

Automated HPLC analyses of drugs of abuse via direct injection of biological fluids followed by simultaneous solid-phase extraction and derivatization with fluorescence detection.

An automated system is described for the simultaneous extraction and derivatization of nucleophilic compounds from various biological media. The method includes the use of a solid-phase reagent containing a 9-fluorenylacetate activated ester. The reagent is based on a controlled pore, polystyrene divinylbenzene support prepared through a silica template procedure. An X-Y-Z robotic arm equipped with a needle is used in conjunction with a syringe pump for aspirating and dispensing samples and standards into the HPLC system. A precolumn cartridge containing the solid-phase reagent is put on-line in place of the fixed-volume injection loop. Injections of biological fluids such as urine or plasma with minimal sample treatment and handling are made directly into this reactor. The analytes are derivatized as they are extracted, allowing virtually unlimited sample volumes to be injected. The polymeric cartridge can be used for up to 100 injections without accruing unacceptable reductions in sensitivity. A detection limit of 500 p.p.t. (parts per trillion) of amphetamine in urine was achieved with this system.